Eukaryotic genomes are replete with DNA microsatellites in which 1-5 base pairs are tandemly repeated. Microsatellites provide a useful "window" into the molecular mechanisms of genetic stability because these simple elements are hotspots for mutations that change the repeat length. For most simple repeats, tract alterations are typically limited to changes of 1-2 repeat units at a time. These sequences are stabilized by DNA mismatch repair, which corrects pre-mutagenic mispairs. In mismatch repair-deficient cells, the mutation frequency is greatly elevated at many microsatellites throughout the genome. Trinucleotide repeats (TNRs) exhibit very different genetic behavior. TNR expansions tend to be much longer and more frequents, once a critical threshold length has been achieved. These large expansions are not subject to mismatch repair because the large hairpin intermediates cannot be corrected. TNR alterations are highly localized; multiple TNRs have not been observed to undergo simultaneous, large alterations. These and other facts suggest that TNRs are subject to unique mutational mechanisms. The biomedical relevance of TNR instability stems from the fact that at least 14 human inherited diseases are caused by TNR expansions. TNR length in the androgen receptor gene is also a key risk factor in prostate cancer. This proposal focuses on the mechanisms of TNR instability in yeast and human cells. Genetic assays have been developed that allow specific identification of cells harboring altered TNR tracts. These assays allow analysis of the important DNA elements, such as thresholds, that govern TNR alterations. A genetic approach also allows isolation of the genes that control TNR instability.